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STUDIES ON SOIL STREPTOMYCETES FROM JORDAN I. SAADOUN and E AL-MOMANI Department of Biological Sciences, Jordan University of Science & Technology, Irbid 22110, Jordan
Code Number: AC97007
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ABSTRACT. An investigation was carried out to evaluate the distribution of soil streptomycetes in different locations in Jordan. Based on sporophore morphology, 339 strains were identified as Streptomyces and 22 strains as Streptoverticillium. Cultural and morphological characteristics and antibiotic activity of the strains are discussed. Streptomycetes are widely distributed in nature, especially in soil. In addition the organisms are among the most important producer of antibiotic and other secondary metabolites.
Data concerning the distribution of members of streptomycetes in Jordanian soils are lacking. However, the occurrence of these organisms in cultivated soils of several plant nurseries in Jordan was previously reported by Abussaud & Saadoun (1988) and Saadoun & Al-Momani (1997).
In the present paper strains belonging to the genera Streptomyces and Streptoverticillium, obtained from 45 soil samples collected from 9 different locations in Jordan are investigated. MATERIALS and METHODS Sampling. Five samples from each location (Fig. 1) were collected during summer 1996. These were taken with an auger up to 10cm depth, after removing approximately 3 cm of the soil surface. Samples were placed in polyethylene bags, closed tightly and stored in a refrigerator.
Characterisation of the isolates. Streptomyces and Streptoverticillium colonies were characterised morphologically and physiologically following the directions given by the International Streptomyces project (ISP) (Shirling & Gottlieb, 1966). General morphology was determined on oatmeal agar plates, incubated in the dark at 27 C for 21 dd, by direct light microscopy examination of the surface of crosshatched cultures. Colours were determined according to Prauser (1964) and isolates were grouped as proposed by Nonomura (1974). Antimicrobial activity. This was tested by plate diffusion method against Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Candida albicans. Isolates were grown on oatmeal agar for 14 dd; discs (9mm in diameter) were cut and placed on nutrient agar seeded with the test organisms and incubated at 27 C. Inhibition zones were checked after 24hrs. RESULTS and DISCUSSION The average microbial count ranged between 10^6 to 10^9 and 10^4 to 10^6 CFU/gm dry soil for bacteria and streptomycetes respectively. A total of 361 streptomycetes were recovered from the samples, 339 isolates were identified as belonging to the genera Streptomyces (Table 1) and 22 to Streptoverticillium (Table 4). Streptomyces isolates were distributed into seven series according to the colour of their mature sporulated aerial mycelium (Table 1). Most of these isolates were recovered from Umm Qays (57) and Umm al Jimal (52). Members of the grey series were found to represent 36% of the total number of the isolates; however, the lowest occurrence was for the violet series (5%). Table 1. Distribution of Streptomyces strains isolated from different sites (see Figure 1).
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LOCATION COLOUR SERIES TOTAL
White Grey Yellow Red Blue Green Violet (%)
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1. Umm Qays 7 22 4 10 6 4 4 57 (17)
2. Ash Shuna 2 8 1 4 5 2 1 23 (7)
3. AI Buwaydah 8 15 7 1 1 6 3 41 (12)
4. Umm al Jimal 16 20 7 1 0 8 0 52 (15)
5. Ajlun 6 18 4 7 5 5 3 48 (14)
6. Deir Alia 3 10 2 7 4 4 1 2 (9)
7. Suwaymah 2 4 2 3 3 3 2 19 (6)
8. Wadi-Musa 5 11 2 6 3 5 2 34 (10)
9. Maan 6 15 4 1 2 6 0 34 (10)
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TOTAL 55 123 33 40 29 43 16 339
(%) (16) (36) (10) (12) (8.5) (12.5) (5) (100)
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Msameh (1992) in his study on distribution and antibiotic activity of Streptomyces flora in Jordan reported that 33.2% of the organisms were isolated from the desert. White and grey colour series showed the highest percentage of occurrence (43.6% and 28.3%, respectively). In the present study, AI Buwaydah, Umm al Jimal and Mann represent the desert areas in Jordan and 37% of the Streptomyces isolates were recovered from these locations. Strains of the grey and white colour series had also the highest percentage of occurrence among the total isolates (36% and 16%, respectively). Data on the highest occurrence of isolates of the grey series are in agreement with those obtained by Hamdi et al. (1980), Abussaud & Saadoun (1988), Msameh (1992) and Saadoun & Al-Momani (1997). Of the 339 Streptomyces isolates, 129 produce melanin, 109 show distinctive reverse side pigment and 68 produce soluble pigments (Table 2). Table 2. Cultural and morphological characteristics of the Streptomyces isolates.
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COLOUR SERIES TOTAL
CHARACTER White Grey Yellow Red Blue Green Violet (%)
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Pigment production:
Melanin 13 46 16 14 27 11 2 129 (42)
Reverse colour 15 31 11 38 1 9 4 109 (36)
Soluble colour 11 18 16 1 1 19 2 68 (22)
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TOTAL 39 95 43 53 29 39 8 306
(%) (13) (31) (14) (17) (9.5) (13) (2.5) (100)
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Sporophore morphology:
Straight 30 19 3 8 7 7 2 76 (22)
Spiral 12 84 10 26 17 23 10 182 (54)
Flexuous 13 14 15 3 4 11 4 64 (19)
Retinaculum 0 6 5 3 1 2 0 17 (5)
apertum
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TOTAL 55 123 33 40 29 43 16 339
(%) (16) (36) (10) (12) (8.5) (12.5) (5) (100)
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With reference to the morphology of spore bearing hyphae most isolates show spiral (54%) sporophores (Table 2) as had been previously reported by Hamdi et al. (1980), Coelho & Drozdowicz (1979), Abussaud & Saadoun (1988) Msameh (1992) and Saadoun & Al-Momani (1997). Antibacterial and antifungal activity of the Streptomyces isolates against the tested bacteria and Candida albicans is shown in Table 3 Table 3. Antibacterial and antifungal activity of the Streptomyces isolates (percentages in brackets).
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COLOUR SERIES TOTAL
ANTIBIOSIS White Grey Yellow Red Blue Green Violet
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Isolates 55 123 40 29 43 33 16 339
Active 53 110 36 22 40 30 14 305
strains (96) (89) (90) (76) (93) (91) (87.5) (90)
S. aureus 18 60 3 2 21 10 3 117
(34) (54.5) (8) (9) (52.5) (33) (21) (39)
B. subtilis 18 41 2 20 14 16 5 116
(34) (37) (6) (91) (35) (53) (36) (36)
E. coli 2 12 0 2 0 3 0 19
(4) (11) (9) (10) (6)
C. albicans 15 28 0 2 0 8 0 53
(28) (25.5) (9) (27) (17)
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Ninety percent of the isolates were active against one or more of the test organisms, however the percentage of active isolates varies within each colour series, tending to be higher in the white and blue series and lower in the red one. Data show that most colour series had almost an equal antibiotic activity. The activity of the different isolates was almost equal against S. aureus (39%) and B. subtilis (38%). The activity against E. coli and C. albicans was also similar (19% and 17%, respectively). Similarly Msameh (1992) reported that the activity of the different Streptomyces isolates against B. cereus and S. aureus was also the same (33.9% and 34.9%, respectively), while 4.6% and 16.7% were active against E. coli and C. albicans. Morphological, cultural and antibiosis characteristics of the 22 isolates attributed to the genus Streptoverticillium (Locci et al., 1969; Nonomura, 1974) are shown in Table 4. Table 4. Characteristics of the Streptoverticillium strains. All isolates utilise arabinose, fructose, xylose (except No.8), mannitol (exc. No.15), rhamnose (exc. No.10 and sucrose (exc. Nos.1 and 4) and are active against S. aureus (exc. Nos.3 and 15). (AM: aerial mycelium colour, Bs: B. subtilis, BV: biverticillate, Ca: Candida albicans, Ec: E. coli, Ge: green, Gy: grey, IN: inositol, ME: melanin pigment, MV: monoverticillate, RC: reverse colour; RA: raffinose, SC: spore chain, SP: soluble pigment, Wh: white).
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STRAIN CULTURAL AND SUGAR ANTIBIOSIS
No. MORPHOLOGICAL CHARACTERS UTILISATION
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AM ME RC SP SC IN RA Ec Bs Ca
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1 Gy - + + MV + + + + +
2 Gy - + + BV + + + + +
3 Gy - + - BV + + - - -
4 Gy - + - BV + - - - -
5 Gy - - - MV + + - - -
6 & 7 Gy - - - BV + + - - -
8 & 9 Gy - - - BV + + + + +
10,11&12 Red - + - BV - - - - -
13 Red - - - MV - - - - -
14 Red - - - BV +/- - + + +
15 Red + + + BV + + + + +
16 Red - + - BV - + - + +
17 Blue + - - BV + + + + +
18 Ge - - - BV - - + + +
19 Ge - - - BV - - - - -
20 Ge - - - BV + + + + +
21 Ge - - - MV - - + + +
22 Wh - + + BV - - - - -
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Streptoverticillium strains are grouped, according to the colour of the aerial mycelium, as grey (9), red (7), green (4), blue and white (1 each). Eighteen strains are characterised by biverticillate and 4 by monoverticillate sporophores (see however, Locci & Schofield, 1989). Two strains are melanin positive, 9 show reverse colour and 4 produce soluble pigments. With reference to cultural and morphological characters, these strains do not fit into Nonomuras key (1974). As regards antibiotic activity, ten strains inhibit all tested organisms. ACKNOWLEDGEMENTS. This research was financed by a grant No. 24/96 from the Deanship of Scientific Research at Jordan University of Science and Technology. The technical assistance of Maher Obiedat is greatly appreciated. REFERENCES Abussaud, M.J & I.M. Saadoun (1988). Isolation, characterization and taxo- nomy of Streptomyces sp. isolated from Jordanian soils and antagonistic to Agrobacterium tumefaciens. Egypt. J. Microbiol., 23:597-609 Coelho, R.R. & A. Drozdowicz (1979). The occurrence of actinomycetes in cerrado soil in Brazil. Rev. Ecol. Biol. Sci., 15:459-474 Hamdi, Y.A., D. Ahmed &A.M. Al-Tai (1980) Genera and species of actinomycetes isolated from Iraqi soils. Egypt. J. Microbiol., 15:7-22 Kuster, E. & S.T. Williams (1964). Selection of media for isolation of streptomycetes, Nature, 202:928-929 Locci, R. & G. Schofield (1989). Genus Streptoverticillium Baldacci 1958, 15, emend. mut. char. Baldacci, Farina and Locci 1966, 168^AL. In S.T. Williams, R.E.Sharpe & J.G.Holt (eds.) Bergey's Manual of Systematic Bacteriology. Williams & Wilkins, Baltimore, Vol. 4, pp. 2492-2504 Locci, R., E. Baldacci & B.P. Baldan (1969). The genus Streptoverticillium. A taxonomic study. Giornale di Microbiologia, 17:1-60 Msameh, Y.M. (1992). Streptomyces in Jordan; distribution and antibiotic activity. M.S. Thesis, Department of Biology, Yarmouk University, Irbid, Jordan Nonomura, H. (1974). Key for classification and identification of 485 species of the streptomycetes included in the ISP. J. Ferm. Tech., 52:78-92 Prauser, H. (1964). Aptness and application of colour for exact description of colours of Streptomyces. Zeitsch. Allgem. Mikrobiol., 4:95-98 Saadoun, I. & F. Al-Momani (1997). Streptomycetes from Jordan soils active against Agrobacterium tumefaciens. Actinomycetes, 8:29-36 Shirling, E.B. & D. Gottlieb (1966). Methods for characterization of Streptomyces species. Int. J. Syst. Bacteriol., 16: 313-340. Copyright 1997 C.E.T.A., The International Centre for Theoretical and Applied Ecology, Gorizia The following images related to this document are available:Line drawing images[ac97007a.gif] |
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